Enhancement of the endocannabinoid system through monoacylglycerol lipase inhibition relieves migraine-associated pain in mice

Abstract

Background: Migraine affects over 1 billion people worldwide and is a leading cause of disability. Targeting the cannabinoid system offers a promising approach for pain and migraine relief. This study evaluated a novel monoacylglycerol lipase (MAGL) inhibitor to prolong endocannabinoid action in acute and chronic mouse models of migraine. It also examined MAGL and cannabinoid receptor 1 (CB₁) mRNA expression in key head pain-processing regions.

Methods: C57BL6/J male and female mice received the human migraine trigger nitroglycerin (NTG) acutely or every other day for 9 days. Allodynia was assessed by von Frey hair stimulation of the periorbital area. A single dose of MAGL inhibitor (ABD-1970) was tested in acute and chronic NTG models. Additionally, ABD-1970 was given daily for 5 days to assess tolerance. In situ hybridization measured transcript expression of MAGL, CB₁, and neuronal marker Rbfox3 in trigeminal ganglia (TG) and trigeminal nucleus caudalis (TNC).

Results: A single injection of ABD-1970 blocked cephalic allodynia induced by acute NTG. ABD-1970 also blocked chronic allodynia established by chronic intermittent NTG. Repeated administration did not induce tolerance, and ABD-1970 continued to block NTG-induced allodynia after 5 days of administration. There was high expression of MAGL and CB₁ in the TG and TNC, present in Rbfox3 positive and negative cells.

Conclusion: MAGL inhibitor effectively blocked acute and chronic migraine-associated pain, likely through prolonged endocannabinoid action. This effect may be mediated through action at peripheral or central sites considering the high MAGL and CB₁ expression in the TG and TNC, respectively. The endocannabinoid system appears to modulate migraine mechanisms, and MAGL may be a promising target for this disorder.

Keywords: Endocannabinoid; Headache; RNAScope; Trigeminovascular pain.

Fig. 1

Acute nitroglycerin (NTG)-induced cephalic allodynia is blocked by acute MAGL inhibition in male and female C57BL6/J mice. (A) Schematic of paradigm includes measuring baseline cephalic thresholds, injecting ABD-1970 (10 mg/kg PO), methylcellulose vehicle control (PO), or olcegepant (1 mg/kg IP) 4 h before von Frey testing (teal circle), injecting nitroglycerin (10 mg/kg IP) or saline vehicle 2 h before testing (open circle), and injecting sumatriptan (0.6 mg/kg IP) 45 min before testing (filled in orange circle). (B) Basal responses and (C) post-drug test responses are shown (n = 8/group). 1-way ANOVA analysis, significant effect of treatment (p < 0.001). Holm-Sidak multiple comparisons: **p < 0.01, ***p < 0.001 relative to NTG-Veh, ###p < 0.001 relative to Veh-Veh. Female animals indicated with open circles. All data are presented as mean ± SEM

Fig. 2

Chronic NTG-induced cephalic allodynia is blocked by acute MAGL inhibition in male and female C57BL6/J mice. (A) Schematic of paradigm includes animals receiving vehicle or NTG (10 mg/kg IP) every other day for 9 days (open circle), with baseline mechanical responses measured on days 1, 5, 9, and 10. Animals were injected with Veh, ABD-1970, olcegepant, or SNC80 and tested (filled in teal circle). (B) Chronic NTG led to a decrease in mechanical threshold that lasted at least ~ 24–48 h after last injection. (n = 8/group). 2-way repeated measures ANOVA analysis, significant effect of time, treatment, and interaction (p < 0.001). Holm-Sidak multiple comparisons: ##p < 0.01, ###p < 0.001 relative to Veh-Veh. (C) Post-drug responses after treatment with ABD-1970 (10 mg/kg PO), methylcellulose vehicle control (PO) or olcegepant (1 mg/kg IP) 4 h before testing, and DOR agonist SNC80 (10 mg/kg IP) or saline vehicle (IP) 45 min before testing post-drug thresholds (n = 8/group). 1-way ANOVA analysis, significant effect of treatment (p < 0.001). Holm-Sidak multiple comparisons: *p < 0.05, **p < 0.01 relative to NTG-Veh, ###p < 0.001 relative to Veh-Veh. Female animals indicated with open circles. All data are presented as mean ± SEM

Fig. 3

Chronic MAGL inhibition caused no change in baseline mechanical threshold and continued to block acute migraine-associated pain in male and female C57BL6/J mice. (A) Schematic of paradigm– mice were treated with chronic MAGL inhibitor (ABD-1970 (10 mg/kg, 5x QD, PO) or methylcellulose vehicle control (PO)) every day for four days (filled in teal circle). On day 5 baselines were taken, and then animals were injected with Veh, ABD-1970, sumatriptan, or olcegepant before taking post-drug thresholds (open circle). (B) On day 5, baselines were taken (n = 8/group). 2-way repeated measures ANOVA analysis showed no significant effect of time (p = 0.8225), treatment (p = 0.2736) or interaction (p = 0.2826) after chronic MAGL inhibition. (C) Then mice were injected with ABD-1970 (10 mg/kg PO), methylcellulose vehicle control (PO) or olcegepant (1 mg/kg IP) 4 h before testing. NTG or saline vehicle (10 mg/kg IP) was administered 2 h before testing, and sumatriptan (0.6 mg/kg IP) 45 min before testing post-drug thresholds (n = 8/group). 1-way ANOVA analysis, significant effect of treatment (p < 0.001). Holm-Sidak multiple comparisons: *p < 0.05, **p < 0.01, ***p < 0.001 relative to NTG-Veh, ##p < 0.01 relative to Veh-Veh. Female animals indicated with open circles. All data are presented as mean ± SEM

Fig. 4

MAGL is highly expressed in the TG and TNC, with higher expression in Rbfox3/NeuN expressing cells. (A) Representative images of fluorescent in situ hybridization. Chevron indicates a cell expressing MAGL and Rbfox3, arrow indicates a cell expressing CB₁ and Rbfox3 transcripts. Percentage of total cells expressing transcripts stained with DAPI in (B) TG and (C) TNC. Percentage of total Rbfox3 + cells expressing MAGL and/or CB₁ transcript in the (C) TG and (E) TNC. Percentage of total Rbfox3 negative cells expressing MAGL and/or CB₁ transcript in the (D) TG and (F) TNC. Data are the average from 5 mice, at least four 20x images/mouse were quantified using HALO software. Data represented as mean ± SEM. Females are represented with clear symbols